JP6834449B2 - Dispersion method and disperser of magnetic particles - Google Patents

Dispersion method and disperser of magnetic particles Download PDF

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JP6834449B2
JP6834449B2 JP2016242696A JP2016242696A JP6834449B2 JP 6834449 B2 JP6834449 B2 JP 6834449B2 JP 2016242696 A JP2016242696 A JP 2016242696A JP 2016242696 A JP2016242696 A JP 2016242696A JP 6834449 B2 JP6834449 B2 JP 6834449B2
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哲也 甲斐
哲也 甲斐
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本発明は、磁性粒子を含む凍結乾燥試薬の製造に適した、磁性粒子の分散方法及び分散装置に関するものである。 The present invention relates to a method for dispersing magnetic particles and a dispersion device suitable for producing a freeze-drying reagent containing magnetic particles.

溶液中に磁性粒子を分散して反応させる凍結乾燥試薬の製造は、磁性粒子を含む懸濁液を調製したのち所定量の懸濁液を容器に分注し、凍結工程、乾燥工程を経る方法が一般的である。溶液に比べて比重の大きな磁性粒子を用いる場合は分注後、凍結工程に移動するまでに容器内で磁性粒子が自然沈降する。磁性粒子の分注から凍結工程への移動に長時間を要する製造設備においては磁性粒子の大部分が容器底部に沈降することもあり、この状態で凍結乾燥すると磁性粒子が製剤の底部に偏在する。 The production of a freeze-drying reagent in which magnetic particles are dispersed and reacted in a solution is a method in which a suspension containing magnetic particles is prepared, a predetermined amount of the suspension is dispensed into a container, and a freezing step and a drying step are performed. Is common. When magnetic particles having a higher specific gravity than the solution are used, the magnetic particles spontaneously settle in the container after dispensing and before moving to the freezing step. In a manufacturing facility where it takes a long time to move the magnetic particles from the dispensing to the freezing process, most of the magnetic particles may settle on the bottom of the container, and if freeze-dried in this state, the magnetic particles are unevenly distributed on the bottom of the preparation. ..

一方、凍結乾燥試薬を保存する容器が測定容器を兼ねる測定法において、前記磁性粒子が容器底部に偏在した凍結乾燥試薬を用いると、測定装置上で凍結乾燥試薬を溶解後、所定の攪拌時間内に磁性粒子の十分な分散が達成されず、測定値が変動するという問題が生じる。そこで、磁性粒子を含む溶液を、凍結乾燥の凍結工程に導入する直前に短時間で均一に分散する方法が望まれる。 On the other hand, in the measurement method in which the container for storing the lyophilization reagent also serves as the measurement container, if the lyophilization reagent in which the magnetic particles are unevenly distributed at the bottom of the container is used, the lyophilization reagent is dissolved on the measuring device and then within a predetermined stirring time. However, sufficient dispersion of the magnetic particles is not achieved, and there arises a problem that the measured value fluctuates. Therefore, a method of uniformly dispersing the solution containing the magnetic particles in a short time immediately before introducing it into the freezing step of freeze-drying is desired.

磁性粒子の分散方法は分析法の分野においては、例えば特許文献１に示されているように攪拌翼を容器内に投入し、磁性粒子を溶液中に分散する方法があるが、攪拌翼の剪断応力により磁性粒子表面が損傷を受ける、さらには撹拌翼による他容器内へのキャリーオーバーが生じる、等の課題をもつ。また、特許文献２は磁石を用いて磁性粒子を容器内壁面に集磁したのち、容器を往復揺動して非接触で磁性粒子を溶液中に分散する方法を開示しているが、容器内壁面に強く集磁するために磁性粒子同士が凝集する、又は集磁面に付着する可能性がある。これらを均一に分散するには攪拌時間を長くする、又は攪拌強度を上げる等の工夫が必要になる。したがって、特許文献１又は２のいずれの方法を用いても磁性粒子にとって過酷な攪拌条件となり、磁性粒子を含む凍結乾燥試薬の製造に適した分散方法とはいえない。 In the field of analytical methods, as a method of dispersing magnetic particles, for example, as shown in Patent Document 1, there is a method of putting a stirring blade into a container and dispersing the magnetic particles in a solution, but shearing of the stirring blade The surface of the magnetic particles is damaged by the stress, and the stirring blade causes carryover into other containers. Further, Patent Document 2 discloses a method of collecting magnetic particles on the inner wall surface of a container using a magnet and then swinging the container back and forth to disperse the magnetic particles in a solution in a non-contact manner. Since the magnetic particles are strongly focused on the wall surface, the magnetic particles may aggregate or adhere to the magnetic collecting surface. In order to disperse these uniformly, it is necessary to take measures such as lengthening the stirring time or increasing the stirring strength. Therefore, using either method of Patent Document 1 or 2 results in severe stirring conditions for the magnetic particles, and it cannot be said that the dispersion method is suitable for producing a freeze-drying reagent containing the magnetic particles.

特開２０１０−１０１６７７号公報JP-A-2010-101677 特開２０１５−１４８５１０号公報JP 2015-148510

容器に収容された磁性粒子を含む凍結乾燥試薬を製造するにあたり、凍結工程の直前において磁性粒子を非接触かつ短時間で均一に分散する方法及びそれを用いた分散装置を提供することを目的とする。 In producing a freeze-drying reagent containing magnetic particles contained in a container, it is an object of the present invention to provide a method for uniformly dispersing magnetic particles in a short time without contact immediately before the freezing step, and a dispersion device using the same. To do.

本発明者らは、鋭意検討を行った結果、磁性粒子と溶液が入った容器の外部から磁界を作用させて、磁性粒子を溶液相と気相の界面に沿った位置に集磁し、容器を水平方向に振動させることにより、磁性粒子を非接触かつ短時間で均一に分散させる方法を見出し、本発明を完成するに至った。 As a result of diligent studies, the present inventors applied a magnetic field from the outside of the container containing the magnetic particles and the solution to collect the magnetic particles at a position along the interface between the solution phase and the gas phase, and the container. The present invention has been completed by finding a method for uniformly dispersing magnetic particles in a short time without contact by vibrating the magnetic particles in the horizontal direction.

前記課題を解決し、目的を達成するためになされた本発明は、以下の発明を包含する。 The present invention made to solve the above problems and achieve the object includes the following inventions.

すなわち、本発明の第一の態様は、
磁性粒子と溶液が入った容器の外部から磁界を作用させて、磁性粒子を溶液相と気相の界面に沿った位置に集磁し、容器を水平方向に振動させる、磁性粒子の分散方法である。磁性粒子は、前記界面の面積の９０％以上となるように広がっていることが好ましい。
また、前記容器を水平方向に振動させると同時に又は振動させる前に、前記磁界を解除又は減衰させることが好ましい。 That is, the first aspect of the present invention is
A magnetic particle dispersion method in which a magnetic field is applied from the outside of a container containing magnetic particles and a solution to collect the magnetic particles at a position along the interface between the solution phase and the gas phase and vibrate the container in the horizontal direction. is there. The magnetic particles are preferably spread so as to be 90% or more of the area of the interface.
Further, it is preferable to release or attenuate the magnetic field at the same time as or before vibrating the container in the horizontal direction.

次に、本発明の第二の態様は、
磁性粒子と溶液が入っている容器の外部から磁界を作用させて前記磁性粒子を溶液相と気相の界面に沿った位置に集磁する集磁手段と、
前記容器を水平方向に振動させる攪拌手段と、
を備えた分散装置である。
また、前記容器は一次元方向又は二次元方向に複数個配列されていても良好な分散が可能である。 Next, the second aspect of the present invention is
A magnetic collecting means for collecting the magnetic particles at a position along the interface between the solution phase and the gas phase by applying a magnetic field from the outside of the container containing the magnetic particles and the solution.
A stirring means for vibrating the container in the horizontal direction and
It is a dispersive device equipped with.
Further, even if a plurality of the containers are arranged in the one-dimensional direction or the two-dimensional direction, good dispersion is possible.

本発明により、溶液と共に容器に収められている磁性粒子の分散を行う際に、容器の外部から磁界を作用させて容器内の磁性粒子を溶液相と気相の界面に沿った位置に集磁し、容器を水平方向に振動させることにより、磁性粒子を非接触かつ短時間で均一に分散することができる。 According to the present invention, when the magnetic particles contained in the container are dispersed together with the solution, a magnetic field is applied from the outside of the container to collect the magnetic particles in the container at a position along the interface between the solution phase and the gas phase. By vibrating the container in the horizontal direction, the magnetic particles can be uniformly dispersed in a short time without contact.

本発明の略円筒形容器に適用した分散装置の一例を示す正面図である。It is a front view which shows an example of the dispersion apparatus applied to the substantially cylindrical container of this invention. 本発明の略円筒形容器に適用した分散装置の一例を示す平面図である。It is a top view which shows an example of the dispersion apparatus applied to the substantially cylindrical container of this invention. 図１又は２に示す分散装置を用いた磁性粒子の集磁完了時の容器の断面図である。It is sectional drawing of the container at the time of completion of magnetic collection of magnetic particles using the disperser shown in FIG. 1 or 2. 本発明の９６穴マイクロタイタープレートに適用した分散装置の一例を示す正面図である。It is a front view which shows an example of the dispersion apparatus applied to the 96-hole microtiter plate of this invention. 本発明の９６穴マイクロタイタープレートに適用した分散装置の一例を示す平面図である。It is a top view which shows an example of the dispersion apparatus applied to the 96-hole microtiter plate of this invention. 図４又は５に示す分散装置を用いた磁性粒子の集磁完了時の９６穴マイクロタイタープレートの断面図である。It is sectional drawing of the 96-hole microtiter plate at the time of completion of magnetic collection of magnetic particles using the disperser shown in FIG. 4 or 5.

以下に本発明を更に詳細に説明する。 The present invention will be described in more detail below.

本発明で用いられる、磁界を作用させる手段としての磁石は特に限定はなく、永久磁石又は電磁石が制限なく使用できる。永久磁石はアルニコ磁石、ＫＳ鋼、ＭＫ鋼、フェライト磁石、サマリウムコバルト磁石、ネオジム磁石等が例示できる。電磁石は直流電磁石又は交流電磁石でも構わない。永久磁石の場合は硬質素材の固定部材に固定し、容器と磁石との距離を、前記固定部材を移動して調節する方法が例示される。硬質素材としては、ポリエチレン、ポリプロピレン、ポリスチレン、ポリアクリル酸、ポリ塩化ビニル、ポリ塩化ビニリデンなどのプラスチック素材やアルミニウム、銅、真鍮、ステンレスなどの非磁性金属素材や木材を使用することができる。 The magnet used in the present invention as a means for applying a magnetic field is not particularly limited, and a permanent magnet or an electromagnet can be used without limitation. Examples of permanent magnets include alnico magnets, KS steel, MK steel, ferrite magnets, samarium-cobalt magnets, neodymium magnets, and the like. The electromagnet may be a DC electromagnet or an AC electromagnet. In the case of a permanent magnet, a method of fixing to a fixing member made of a hard material and adjusting the distance between the container and the magnet by moving the fixing member is exemplified. As the hard material, plastic materials such as polyethylene, polypropylene, polystyrene, polyacrylic acid, polyvinyl chloride, and polyvinylidene chloride, non-magnetic metal materials such as aluminum, copper, brass, and stainless steel, and wood can be used.

本発明の、磁性粒子と溶液が入った容器の外部から磁界を作用させて集磁するとは、容器の上方から容器面に対して磁石を近づけて溶液相の上端（溶液相と気相の界面に沿った位置）に集磁する方法が例示される。磁界の作用は、固定部材に固定した磁石を昇降機にて下降させる、又は磁性粒子と溶液が入っている容器を昇降機にて上昇させる等の方法により行うことができるが、操作性や容器からの液飛散の危険性を考慮すると、固定部材に固定した磁石を昇降機にて下降させる方法が好ましい。電磁石を用いる場合には電流のＯＮ／ＯＦＦにて磁力を制御できるので電磁石を固定し、前記昇降機を省略することもできる。 In the present invention, magnetizing by applying a magnetic field from the outside of a container containing magnetic particles and a solution means that a magnet is brought close to the container surface from above the container and the upper end of the solution phase (the interface between the solution phase and the gas phase). A method of concentrating magnetism at a position along the above is illustrated. The action of the magnetic field can be performed by a method such as lowering a magnet fixed to a fixing member with an elevator or raising a container containing magnetic particles and a solution with an elevator, but the operability and the container can be used. Considering the risk of liquid scattering, a method of lowering a magnet fixed to a fixing member with an elevator is preferable. When an electromagnet is used, the magnetic force can be controlled by turning the current on and off, so that the electromagnet can be fixed and the elevator can be omitted.

本発明の、磁性粒子を溶液相と気相の界面に沿った位置に集磁するとは、容器の上方から作用する磁界の強度を調節して磁性粒子が溶液相と気相の界面に沿うように集磁する、すなわち、容器内壁面の一部に集磁するのではなく、略円筒形の容器では溶液相と気相の界面面積と同等に広く集磁することを意味する。具体的には、溶液相と気相の界面面積の９０％以上となるように磁性粒子が広がっていることが好ましく、９５％以上となっていることがより好ましい。磁性粒子が溶液相の上端の位置に集磁されているため、磁界解除後に磁性粒子が溶液相の上端から自然沈降するのと同期して水平方向の振動が行えるため、短時間で容易に溶液相内の均一分散が達成される。 In the present invention, when the magnetic particles are focused at a position along the interface between the solution phase and the gas phase, the strength of the magnetic field acting from above the container is adjusted so that the magnetic particles are along the interface between the solution phase and the gas phase. In other words, instead of concentrating on a part of the inner wall surface of the container, it means that in a substantially cylindrical container, the magnetism is as wide as the interface area between the solution phase and the gas phase. Specifically, the magnetic particles are preferably spread so as to be 90% or more of the interface area between the solution phase and the gas phase, and more preferably 95% or more. Since the magnetic particles are concentrated at the upper end of the solution phase, horizontal vibration can be performed in synchronization with the natural sedimentation of the magnetic particles from the upper end of the solution phase after the magnetic field is released, so the solution can be easily prepared in a short time. Uniform dispersion within the phase is achieved.

本発明で用いられる磁石と容器の接近距離は、磁石の磁力、磁性粒子の磁気応答性、溶液の密度や粘度などから決定される。磁石との距離が遠すぎる場合には磁性粒子が気相と溶液相の界面に沿った位置に集まらず、容器の底面や側面などに残存するため分散効率が落ちる。その反対に磁石との距離が近すぎる場合には、溶液中にある磁性粒子が飛び出して磁石自体に捕集されてしまうといった不具合が起こる。接近距離の設定に当たっては、予備的に実験を行って決定することが望ましい。 The close distance between the magnet and the container used in the present invention is determined from the magnetic force of the magnet, the magnetic responsiveness of the magnetic particles, the density and viscosity of the solution, and the like. If the distance from the magnet is too long, the magnetic particles will not collect at the position along the interface between the gas phase and the solution phase and will remain on the bottom surface and side surfaces of the container, resulting in a decrease in dispersion efficiency. On the other hand, if the distance to the magnet is too close, the magnetic particles in the solution will pop out and be collected by the magnet itself. When setting the approach distance, it is desirable to make a preliminary experiment to determine it.

本発明で用いられる磁性粒子は、ガラス、金属、セラミックス等の無機物又は高分子ポリマー等の有機物の粒子表面、粒子内部もしくは粒子表面と内部の両方に磁性体を含むものであるが、材質が金属製の微粒子の場合、それ自体を磁性体として用いてもよい。磁性粒子に抗体や牛血清アルブミンなどの蛋白質が結合していてもよく、ポリエチレングリコールやポリプロピレングリコールなどの有機化合物が結合していてもよい。磁性粒子の粒子径は０．１から１００μｍが好ましく、さらには１から１０μｍが好ましい。 The magnetic particles used in the present invention contain magnetic substances on the particle surface of inorganic substances such as glass, metal, ceramics, or organic substances such as high molecular polymer, inside the particles, or both inside and outside the particles, but the material is made of metal. In the case of fine particles, the fine particles themselves may be used as a magnetic material. A protein such as an antibody or bovine serum albumin may be bound to the magnetic particles, or an organic compound such as polyethylene glycol or polypropylene glycol may be bound to the magnetic particles. The particle size of the magnetic particles is preferably 0.1 to 100 μm, more preferably 1 to 10 μm.

本発明に用いられる磁性粒子に含まれる磁性体は、その組成に特に限定はなく、例えば、フェライト、マグネタイト、マグヘマイト等があげられる。磁性粒子の比重は１．０から８．０が好ましく、さらには１．０から６．０がより好ましい。 The composition of the magnetic material contained in the magnetic particles used in the present invention is not particularly limited, and examples thereof include ferrite, magnetite, and maghemite. The specific gravity of the magnetic particles is preferably 1.0 to 8.0, more preferably 1.0 to 6.0.

本発明で用いられる容器の形状は特に限定はなく、略円筒形、略角筒形、又は市販のセラムチューブ等の開口部が１つの容器、９６穴マイクロタイタープレート又は３８４穴マイクロタイタープレート等の二次元方向に複数個配列されている容器、分割型９６穴マイクロタイタープレートのストリップ又は多連型カートリッジ等の一次元方向に複数個配列されている容器であってもよい。 The shape of the container used in the present invention is not particularly limited, and may be a container having a substantially cylindrical shape, a substantially square cylinder shape, or a commercially available serum tube or the like having one opening, a 96-hole microtiter plate, a 384-hole microtiter plate, or the like. It may be a container in which a plurality of containers are arranged in the two-dimensional direction, a container in which a plurality of containers are arranged in the one-dimensional direction such as a strip of a split type 96-hole microtiter plate or a multiple cartridge.

本発明で用いられる容器の大きさは、容器の外部から作用させる磁界の強度に対して溶液相と気相の界面に沿った位置に集磁が可能な容器の大きさであれば制限なく使用できる。 The size of the container used in the present invention is not limited as long as it is the size of a container that can collect magnetism at a position along the interface between the solution phase and the gas phase with respect to the strength of the magnetic field acting from the outside of the container. it can.

本発明で用いられる容器の材質は、一例として、オレフィン系樹脂、スチレン系樹脂、ビニル系樹脂、カーボネート系樹脂、エポキシ系樹脂、アクリル系樹脂、ポリエステル系樹脂、ポリアミド系樹脂といった熱可塑性樹脂又はガラスが挙げられる。ステンレスやアルミなどの非磁性金属、アルミナやジルコニアなどのセラミック、さらには陶器や磁器等であっても特に問題なく使用できる。 The material of the container used in the present invention is, for example, a thermoplastic resin such as an olefin resin, a styrene resin, a vinyl resin, a carbonate resin, an epoxy resin, an acrylic resin, a polyester resin, or a polyamide resin, or glass. Can be mentioned. Non-magnetic metals such as stainless steel and aluminum, ceramics such as alumina and zirconia, and even pottery and porcelain can be used without any problem.

本発明の磁性粒子分散に用いる溶媒としては、水、エタノールやメタノールなどのアルコール類、アセトンやメチルエチルケトンなどのケトン類、ジメチルスルホキシドやＮ，Ｎ−ジメチルホルムアミドなどの非プロトン性極性溶媒、ジエチルエーテル、テトラヒドロフランなどのエーテル類、ベンゼンやトルエンなどの芳香族炭化水素、ヘキサンやヘプタンなどの脂肪族炭化水素を用いることができる。これら溶媒に牛血清アルブミンやコラーゲンペプチドなどの蛋白質、Ｔｒｉｓ−ＨＣｌやＭＥＳなどの緩衝剤、アジ化ナトリウムやプロクリン３００などの防腐剤、スクロースやトレハロースなどの糖、Ｔｗｅｅｎ２０やＴｒｉｔｏｎＸ−１００などの界面活性剤等から一つもしくは二つ以上の成分が含まれていても構わない。 As the solvent used for the dispersion of magnetic particles of the present invention, water, alcohols such as ethanol and methanol, ketones such as acetone and methyl ethyl ketone, aprotic polar solvents such as dimethyl sulfoxide and N, N-dimethylformamide, diethyl ether, etc. Ethers such as tetrahydrofuran, aromatic hydrocarbons such as benzene and toluene, and aliphatic hydrocarbons such as hexane and heptane can be used. These solvents include proteins such as bovine serum albumin and collagen peptides, buffers such as Tris-HCl and MES, preservatives such as sodium azide and procline 300, sugars such as sucrose and trehalose, and surfactants such as Tween 20 and Triton X-100. One or more components may be contained from the agent or the like.

本発明の磁性粒子分散装置は、磁性粒子と溶液が入っている容器の外部から磁界を作用させて前記磁性粒子を溶液相と気相の界面に沿った位置に集磁する集磁手段と、前記容器を水平方向に振動させる攪拌手段と、を備えた分散装置である。 The magnetic particle disperser of the present invention comprises a magnetic collecting means for collecting the magnetic particles at a position along the interface between the solution phase and the gas phase by applying a magnetic field from the outside of the container containing the magnetic particles and the solution. A disperser including a stirring means for vibrating the container in the horizontal direction.

前記磁性粒子集磁手段は、磁性粒子と溶液が入っている容器の上方において磁石を固定した固定部材を昇降可能に設けた集磁手段、又は前記容器の上方において固定部材に固定された磁石に対して前記容器を保持する保持座を昇降可能に設けた集磁手段が例示される。 The magnetic particle magnetic collecting means is a magnetic collecting means provided with a fixing member having a magnet fixed above the container containing magnetic particles and a solution so as to be able to move up and down, or a magnet fixed to the fixing member above the container. On the other hand, a magnetic collecting means in which a holding seat for holding the container can be raised and lowered is exemplified.

前記攪拌手段は、前記容器（溶液相と気相の界面に沿った位置に集磁した磁性粒子を含む）を保持する保持座を水平方向に直線振動及び／又は円振動させる等の単純な攪拌手段が例示される。 The stirring means is a simple stirring such as linear vibration and / or circular vibration of a holding seat holding the container (including magnetic particles collected at a position along the interface between the solution phase and the gas phase) in the horizontal direction. Means are exemplified.

以下、実施例を用いて本発明をさらに詳細に説明するが、これら実施例は本発明を限定するものではない。 Hereinafter, the present invention will be described in more detail with reference to Examples, but these Examples do not limit the present invention.

（実施例１）
図１は本発明の略円筒形容器に適用した分散装置１００の正面図である。図２は本発明の略円筒形容器に適用した分散装置１００の平面図である。表面磁束密度が２，２４３ガウス（Ｇ）のネオジム磁石１０（角型：２００ｍｍ×１５ｍｍ×５ｍｍ、５ｍｍ方向に着磁）１個をステンレス製の固定部材１１に固定し、容器ブロック５２（１６個の容器が搭載可能）に搭載した容器の上方で磁石を昇降可能にする昇降機１３ａに固定した。昇降機１３ａはプレート１２に固定し、プレート１２とともに搬送機１３ｂに沿って攪拌ポジションまで移動可能に設けた。容器は開口径６．２ｍｍ、高さ２０ｍｍの略円筒形容器５１を用いた。容器には磁性粒子７０（粒径２．５μｍ、高分子ポリマーに磁性体を含むタイプ）を、５％牛血清アルブミンを含む希釈液にて０．０８％に調製した懸濁液として５０μＬ分注した。 (Example 1)
FIG. 1 is a front view of the dispersion device 100 applied to the substantially cylindrical container of the present invention. FIG. 2 is a plan view of the dispersion device 100 applied to the substantially cylindrical container of the present invention. One neodymium magnet 10 (square: 200 mm × 15 mm × 5 mm, magnetized in the 5 mm direction) having a surface magnetic flux density of 2,243 gauss (G) is fixed to a stainless steel fixing member 11 and a container block 52 (16 pieces). The magnet was fixed to the elevator 13a that allows the magnet to be raised and lowered above the container mounted on the container. The elevator 13a was fixed to the plate 12 and provided so as to be movable together with the plate 12 to the stirring position along the conveyor 13b. As the container, a substantially cylindrical container 51 having an opening diameter of 6.2 mm and a height of 20 mm was used. In the container, 50 μL of magnetic particles 70 (particle size 2.5 μm, polymer polymer containing magnetic material) was dispensed as a suspension prepared to 0.08% with a diluted solution containing 5% bovine serum albumin. did.

図３は図１又は２に示す分散装置１００を用いた磁性粒子の集磁完了時の容器の断面図である。昇降機１３ａを下降させてネオジム磁石１０を容器の上面から２ｍｍの距離（液面から磁石までの距離は約１９ｍｍ）まで容器の上方から接近させ、２秒間集磁して磁性粒子７０を溶液相と気相の界面に沿って広く（界面面積の１００％程度に）集め、その後、昇降機１３ａを上昇させて磁石１０の磁界を解除した。集磁が完了した容器を前記容器ブロック５２に搭載した状態で、プレート１２に固定した容器ブロック移送手段１４ａにて搬送機１３ｂに沿って攪拌ポジションに移送し、前記容器ブロックを、撹拌機１６を用い、水平方向の直線振動（振動幅２ｍｍ、振動回数２４回／秒、振動時間２秒）により磁性粒子７０を分散した（磁界解除後、振動開始まで４秒）。分散操作が完了した容器ブロックを駆動用シリンダ１５に固定した容器ブロック移送手段１４ｂにて凍結機３０（−４０℃）に移し（振動終了後、凍結機に入るまで５秒）、凍結終了後、乾燥工程を経て磁性粒子の分散性を高めた凍結乾燥製剤を得た。磁性粒子を溶液相と気相の界面に沿った位置に集磁したのち分散する方法は、略円筒形容器底面に沈降した状態から水平方向に振動を加えた分散、又は略円筒形容器の側壁に集磁した状態から振動を加えた分散に比べて短時間に達成され、かつ容器内壁面への磁性粒子の付着が生じないことを確認した。 FIG. 3 is a cross-sectional view of the container when the magnetic particles are collected by using the disperser 100 shown in FIG. 1 or 2. The elevator 13a is lowered to bring the neodymium magnet 10 closer to the container from above the container to a distance of 2 mm (the distance from the liquid surface to the magnet is about 19 mm) from the upper surface of the container, and the magnetic particles 70 are focused on the solution phase for 2 seconds. It was collected widely along the interface of the gas phase (to about 100% of the interface area), and then the elevator 13a was raised to release the magnetic field of the magnet 10. With the container with the completed magnetism mounted on the container block 52, the container block transfer means 14a fixed to the plate 12 transfers the container block to the stirring position along the transporter 13b, and the container block is transferred to the stirrer 16. The magnetic particles 70 were dispersed by linear vibration in the horizontal direction (vibration width 2 mm, vibration frequency 24 times / second, vibration time 2 seconds) (4 seconds from the release of the magnetic field to the start of vibration). The container block for which the dispersion operation has been completed is transferred to the freezer 30 (-40 ° C.) by the container block transfer means 14b fixed to the drive cylinder 15 (5 seconds after the vibration ends until it enters the freezer), and after freezing is completed. A freeze-dried preparation having improved dispersibility of magnetic particles was obtained through a drying step. The method of concentrating the magnetic particles at a position along the interface between the solution phase and the gas phase and then dispersing them is as follows: dispersion by applying vibration in the horizontal direction from the state of being settled on the bottom surface of a substantially cylindrical container, or the side wall of a substantially cylindrical container. It was confirmed that the dispersion was achieved in a shorter time than the dispersion in which the magnetism was applied to the magnet, and that the magnetic particles did not adhere to the inner wall surface of the container.

（実施例２）
図４は本発明の９６穴マイクロタイタープレートに適用した分散装置２００の正面図である。図５は本発明の９６穴マイクロタイタープレートに適用した分散装置２００の平面図である。表面磁束密度が１，８４０ガウス（Ｇ）のネオジム磁石２０（角型：１５０ｍｍ×２０ｍｍ×５ｍｍ、５ｍｍ方向に着磁）５個を各磁石の第２面が相互に接するように結合して（１５０ｍｍ×１００ｍｍ×５ｍｍ）ステンレス製の固定部材２１に固定し、平底９６穴マイクロタイタープレート６１の上方で磁石を昇降可能にする昇降機２３ａに固定した。昇降機２３ａはプレート２２に固定し、プレート２２とともに搬送機２３ｂに沿って攪拌ポジションまで移動可能に設けた。各ウエルには磁性粒子８０（粒径２．５μｍ、高分子ポリマーに磁性体を含むタイプ）を、５％牛血清アルブミンを含む希釈液にて０．０８％に調製した懸濁液として１００μＬ分注した。 (Example 2)
FIG. 4 is a front view of the disperser 200 applied to the 96-hole microtiter plate of the present invention. FIG. 5 is a plan view of the disperser 200 applied to the 96-hole microtiter plate of the present invention. Five neodymium magnets 20 (square: 150 mm x 20 mm x 5 mm, magnetized in the 5 mm direction) with a surface magnetic flux density of 1,840 gauss (G) are combined so that the second surfaces of each magnet are in contact with each other ( It was fixed to a fixing member 21 made of stainless steel (150 mm × 100 mm × 5 mm), and fixed to an elevator 23a that allows the magnet to be elevated above the flat bottom 96-hole microtiter plate 61. The elevator 23a was fixed to the plate 22 and provided so as to be movable together with the plate 22 to the stirring position along the conveyor 23b. In each well, 100 μL of magnetic particles 80 (particle size 2.5 μm, polymer polymer containing magnetic material) was prepared as a suspension prepared to 0.08% with a diluted solution containing 5% bovine serum albumin. Noted.

図６は図４又は５に示す分散装置２００を用いた磁性粒子の集磁完了時の９６穴マイクロタイタープレートの断面図である。昇降機２３ａを下降させてネオジム磁石２０は前記平底９６穴マイクロタイタープレートの上面から９ｍｍの距離（液面から磁石までの距離は約１５ｍｍ）まで容器の上方から接近させ、２秒間集磁して磁性粒子を溶液相と気相の界面に沿って広く（界面の１００％程度に）集め、その後、昇降機２３ａを上昇させ、磁石２０の磁界を解除した。集磁が完了した平底９６穴マイクロタイタープレート６１をプレート２２に固定したマイクロタイタープレート送手段２４ａにて搬送機２３ｂに沿って攪拌ポジションに移送し、前記平底９６穴マイクロタイタープレート６１を、撹拌機２６を用い、水平方向の円振動（水平偏心振とう：振れ幅２ｍｍ、振とう数５００ｒｐｍ、振とう時間５秒）により磁性粒子８０を分散した（磁界解除後、振動開始まで４秒）。分散操作が完了した平底９６穴マイクロタイタープレート６１を駆動用シリンダ２５に固定したマイクロタイタープレート移送手段２４ｂにて凍結機４０（−４０℃）に移し（振動終了後、凍結機に入るまで５秒）、凍結終了後、乾燥工程を経て磁性粒子の分散性を高めた凍結乾燥製剤を得た。磁性粒子を溶液相と気相の界面に沿った位置に集磁したのち分散する方法は、平底９６穴マイクロタイタープレート底面に集磁した状態から振動を加えた分散に比べて短時間に達成され、かつ容器内壁面への磁性粒子の付着が生じないことを確認した。さらに、容器の上方から磁界を作用させることができるため磁石を容器側面に配置する必要がなく、かつ水平方向に振動させる分散方法であるため、分析に多用されている９６穴マイクロタイタープレート等の二次元方向に複数個配列されている形状の容器にも容易に適用することができた。 FIG. 6 is a cross-sectional view of a 96-hole microtiter plate when magnetic particles are collected by using the disperser 200 shown in FIG. 4 or 5. By lowering the elevator 23a, the neodymium magnet 20 is brought close to the top surface of the flat-bottomed 96-hole microtiter plate from above the container to a distance of 9 mm (the distance from the liquid surface to the magnet is about 15 mm), and magnetized for 2 seconds to magnetize. The particles were widely collected along the interface between the solution phase and the gas phase (about 100% of the interface), and then the elevator 23a was raised to release the magnetic field of the magnet 20. The flat-bottomed 96-hole microtiter plate 61 that has been magnetized is transferred to the stirring position along the conveyor 23b by the microtiter plate feeding means 24a fixed to the plate 22, and the flat-bottomed 96-hole microtiter plate 61 is transferred to the stirring position by the stirrer. Using 26, the magnetic particles 80 were dispersed by horizontal circular vibration (horizontal eccentric vibration: swing width 2 mm, shaking number 500 rpm, shaking time 5 seconds) (4 seconds until the start of vibration after the magnetic field was released). The flat-bottomed 96-hole microtiter plate 61 for which the dispersion operation has been completed is transferred to the freezer 40 (-40 ° C.) by the microtiter plate transfer means 24b fixed to the drive cylinder 25 (after the vibration is completed, it takes 5 seconds to enter the freezer. ), After the completion of freezing, a freeze-dried preparation having improved dispersibility of magnetic particles was obtained through a drying step. The method of concentrating the magnetic particles at a position along the interface between the solution phase and the gas phase and then dispersing them is achieved in a shorter time than the dispersion in which the magnetic particles are focused on the bottom surface of the flat-bottomed 96-hole microtiter plate and then vibrated. Moreover, it was confirmed that the magnetic particles did not adhere to the inner wall surface of the container. Further, since a magnetic field can be applied from above the container, it is not necessary to arrange a magnet on the side surface of the container, and since it is a dispersion method that vibrates in the horizontal direction, a 96-hole microtiter plate or the like, which is often used for analysis, is used. It could be easily applied to a container having a shape in which a plurality of containers are arranged in a two-dimensional direction.

また、本発明の分散方法は、凍結乾燥製剤の製造のみならず、前記のように製造して得られた凍結乾燥製剤入り９６穴マイクロタイタープレートを用いて免疫測定する場合においても、免疫反応工程、洗浄工程、酵素反応工程、等の直前に行う磁性粒子の分散方法としても使用することができる。 Further, the dispersion method of the present invention is not only for producing a lyophilized preparation, but also for immunoassay using a 96-well microtiter plate containing a lyophilized preparation obtained as described above. It can also be used as a method for dispersing magnetic particles immediately before a washing step, an enzyme reaction step, or the like.

（比較例１）
実施例１と同様のネオジム磁石、容器ブロック、略円筒形容器、磁性粒子、希釈液を使用した。 (Comparative Example 1)
The same neodymium magnet, container block, substantially cylindrical container, magnetic particles, and diluent as in Example 1 were used.

０．０８％に調製した懸濁液を略円筒形容器に５０μＬ分注した。ネオジム磁石を容器の底面から２０ｍｍの距離まで容器の下方から接近させ、２秒間集磁して磁性粒子を容器の底面に集め、その後ネオジム磁石の磁界を解除した。集磁が完了した容器を前記容器ブロックに搭載した状態で、前記容器ブロックを、撹拌機を用い水平方向の直線振動（振動幅２ｍｍ、振動回数２４回／秒、振動時間２秒）により磁性粒子を分散した（磁界解除後、振動開始まで４秒）。容器底面を確認したところ、磁性粒子の底面付着が確認された。 50 μL of the suspension prepared to 0.08% was dispensed into a substantially cylindrical container. The neodymium magnet was brought close to the bottom surface of the container from the bottom of the container to a distance of 20 mm, and the magnetic particles were collected for 2 seconds on the bottom surface of the container, and then the magnetic field of the neodymium magnet was released. With the magnetically collected container mounted on the container block, the container block is subjected to horizontal linear vibration (vibration width 2 mm, vibration frequency 24 times / second, vibration time 2 seconds) using a stirrer. (4 seconds from the release of the magnetic field to the start of vibration). When the bottom surface of the container was checked, it was confirmed that the magnetic particles adhered to the bottom surface.

（比較例２）
実施例１と同様のネオジム磁石、容器ブロック、略円筒形容器、磁性粒子、希釈液を使用した。 (Comparative Example 2)
The same neodymium magnet, container block, substantially cylindrical container, magnetic particles, and diluent as in Example 1 were used.

０．０８％に調製した懸濁液を略円筒形容器に５０μＬ分注した。ネオジム磁石を容器の側面から２０ｍｍの距離まで容器の片方側面から接近させ、２秒間集磁して磁性粒子を容器の側面に集め、その後ネオジム磁石の磁界を解除した。集磁が完了した容器を前記容器ブロックに搭載した状態で、前記容器ブロックを、撹拌機を用い水平方向の直線振動（振動幅２ｍｍ、振動回数２４回／秒、振動時間２秒）により磁性粒子を分散した（磁界解除後、振動開始まで４秒）。容器側面を確認したところ、磁性粒子の側面付着が確認された。 50 μL of the suspension prepared to 0.08% was dispensed into a substantially cylindrical container. The neodymium magnet was approached from one side of the container to a distance of 20 mm from the side surface of the container, and the magnetic particles were collected for 2 seconds on the side surface of the container, and then the magnetic field of the neodymium magnet was released. With the magnetically collected container mounted on the container block, the container block is subjected to horizontal linear vibration (vibration width 2 mm, vibration frequency 24 times / second, vibration time 2 seconds) using a stirrer. (4 seconds from the release of the magnetic field to the start of vibration). When the side surface of the container was checked, it was confirmed that the magnetic particles adhered to the side surface.

１０、２０： ネオジム磁石
１１、２１： 固定部材
１２、２２： プレート
１３ａ、２３ａ： 昇降機
１３ｂ、２３ｂ： 搬送機
１４ａ、１４ｂ： 容器ブロック移送手段
１５、２５： シリンダ
１６、２６： 撹拌機
２４ａ、２４ｂ： マイクロタイタープレート移送手段
３０、４０： 凍結機
５１： 略円筒形容器
５２： 容器ブロック
６１： 平底９６穴マイクロタイタープレート
７０、８０： 磁性粒子
１００、２００： 分散装置 10, 20: Neodymium magnets 11, 21: Fixing members 12, 22: Plates 13a, 23a: Elevators 13b, 23b: Conveyors 14a, 14b: Container block transfer means 15, 25: Cylinders 16, 26: Stirrers 24a, 24b : Microtiter plate transfer means 30, 40: Freezer 51: Approximately cylindrical container 52: Container block 61: Flat bottom 96-hole microtiter plate 70, 80: Magnetic particles 100, 200: Disperser

Claims (5)

磁性粒子と溶液が入った容器の上方から磁界を作用させて、
前記磁性粒子を溶液相と気相の界面に沿った位置に集磁し、
前記容器を水平方向に振動させる、
磁性粒子の分散方法。 Magnetic field is applied from above the container containing magnetic particles and solution.
The magnetic particles are focused at a position along the interface between the solution phase and the gas phase.
Vibrate the container horizontally,
Dispersion method of magnetic particles. 前記磁性粒子が前記界面の面積の９０％以上となるように広がった状態で集磁することを特徴とする請求項１に記載の分散方法。 The dispersion method according to claim 1, wherein the magnetic particles are focused in a state of being spread so as to be 90% or more of the area of the interface. 前記容器を水平方向に振動させると同時に又は振動させる前に、前記磁界を解除又は減衰させる、請求項１又は２に記載の磁性粒子の分散方法。 The method for dispersing magnetic particles according to claim 1 or 2, wherein the magnetic field is released or attenuated at the same time as or before the container is vibrated in the horizontal direction. 磁性粒子と溶液が入った容器の上方から磁界を作用させて前記磁性粒子を溶液相と気相の界面に沿った位置に集磁する集磁手段と、
前記容器を水平方向に振動させる攪拌手段と、
を備えた分散装置。 A magnetic collecting means for collecting the magnetic particles at a position along the interface between the solution phase and the gas phase by applying a magnetic field from above the container containing the magnetic particles and the solution.
A stirring means for vibrating the container in the horizontal direction and
Dispersing device equipped with. 前記容器が一次元方向又は二次元方向に複数個配列されている、請求項４に記載の分散装置。 The disperser according to claim 4, wherein a plurality of the containers are arranged in a one-dimensional direction or a two-dimensional direction.
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